Limits...
Kinetic transcriptome analysis reveals an essentially intact induction system in a cellulase hyper-producer Trichoderma reesei strain.

Poggi-Parodi D, Bidard F, Pirayre A, Portnoy T, Blugeon C, Seiboth B, Kubicek CP, Le Crom S, Margeot A - Biotechnol Biofuels (2014)

Bottom Line: Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated.The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei.Genes from two specific clusters might be potential targets for such genetic engineering.

View Article: PubMed Central - PubMed

Affiliation: IFP Energies nouvelles, 1-4 avenue de Bois-Préau, 92852 Rueil-Malmaison, France ; Sorbonne Universités, UPMC Univ Paris 06, Institut de Biologie Paris-Seine (IBPS), F-75005 Paris, France.

ABSTRACT

Background: The filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome and transcriptome sequencing) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG 14 and RUT C30. In this follow-up work, we examine how these mutations affect phenotype as regards the transcriptome and cultivation behaviour.

Results: We performed kinetic transcriptome analysis of the NG 14 and RUT C30 strains of early enzyme production induced by lactose using bioreactor cultivations close to an industrial cultivation regime. RUT C30 exhibited both earlier onset of protein production (3 h) and higher steady-state productivity. A rather small number of genes compared to previous studies were regulated (568), most of them being specific to the NG 14 strain (319). Clustering analysis highlighted similar behaviour for some functional categories and allowed us to distinguish between induction-related genes and productivity-related genes. Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting that these clusters contain genes playing a role in strain performance.

Conclusions: This is the first kinetic analysis of a transcriptomic study carried out under conditions approaching industrial ones with two related strains of T. reesei showing distinctive cultivation behaviour. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei. Genes from two specific clusters might be potential targets for such genetic engineering.

No MeSH data available.


Related in: MedlinePlus

Differences in gene expression before induction of protein production (T0) in NG 14 and RUT C30 identified by RNA-seq. The percentage of genes with superior basal level (red bars), inferior basal level (green bars) or equal basal level (grey) is shown for each cluster.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4279801&req=5

Fig4: Differences in gene expression before induction of protein production (T0) in NG 14 and RUT C30 identified by RNA-seq. The percentage of genes with superior basal level (red bars), inferior basal level (green bars) or equal basal level (grey) is shown for each cluster.

Mentions: As we were surprised by the large number of genes regulated in the NG 14 strain, we attempted to explain the apparent loss of up- and down-regulation in the RUT C30 strains. To achieve this, the time 0 references from RUT C30 and NG 14 were compared using RNA-seq experiments (Additional file 6: Table S3). From the 568 differentially expressed genes found using microarrays, we identified by RNA-seq only 23 whose expression was different between the two strains (Table 2), meaning that 95.9% of the differentially expressed genes during the protein production have the same basal level in these strains. The distribution of these genes between each cluster shows that most of them are included in clusters 8 and 9 (Figure 4). Most of these genes are cellulase genes that have a higher basal expression in RUT C30 and that represent 60% of cluster 9 (seven genes) and 10% of cluster 8 (four genes). This may explain the apparent lower induction of these genes in RUT C30 and is compatible with our previously published results positively correlating induction ratios and protein production [34]. However, the small number of genes induced or repressed prior to the lactose induction suggests that expression patterns observed between the two strains indeed reflect different behaviours regarding the lactose induction signal.Table 2


Kinetic transcriptome analysis reveals an essentially intact induction system in a cellulase hyper-producer Trichoderma reesei strain.

Poggi-Parodi D, Bidard F, Pirayre A, Portnoy T, Blugeon C, Seiboth B, Kubicek CP, Le Crom S, Margeot A - Biotechnol Biofuels (2014)

Differences in gene expression before induction of protein production (T0) in NG 14 and RUT C30 identified by RNA-seq. The percentage of genes with superior basal level (red bars), inferior basal level (green bars) or equal basal level (grey) is shown for each cluster.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4279801&req=5

Fig4: Differences in gene expression before induction of protein production (T0) in NG 14 and RUT C30 identified by RNA-seq. The percentage of genes with superior basal level (red bars), inferior basal level (green bars) or equal basal level (grey) is shown for each cluster.
Mentions: As we were surprised by the large number of genes regulated in the NG 14 strain, we attempted to explain the apparent loss of up- and down-regulation in the RUT C30 strains. To achieve this, the time 0 references from RUT C30 and NG 14 were compared using RNA-seq experiments (Additional file 6: Table S3). From the 568 differentially expressed genes found using microarrays, we identified by RNA-seq only 23 whose expression was different between the two strains (Table 2), meaning that 95.9% of the differentially expressed genes during the protein production have the same basal level in these strains. The distribution of these genes between each cluster shows that most of them are included in clusters 8 and 9 (Figure 4). Most of these genes are cellulase genes that have a higher basal expression in RUT C30 and that represent 60% of cluster 9 (seven genes) and 10% of cluster 8 (four genes). This may explain the apparent lower induction of these genes in RUT C30 and is compatible with our previously published results positively correlating induction ratios and protein production [34]. However, the small number of genes induced or repressed prior to the lactose induction suggests that expression patterns observed between the two strains indeed reflect different behaviours regarding the lactose induction signal.Table 2

Bottom Line: Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated.The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei.Genes from two specific clusters might be potential targets for such genetic engineering.

View Article: PubMed Central - PubMed

Affiliation: IFP Energies nouvelles, 1-4 avenue de Bois-Préau, 92852 Rueil-Malmaison, France ; Sorbonne Universités, UPMC Univ Paris 06, Institut de Biologie Paris-Seine (IBPS), F-75005 Paris, France.

ABSTRACT

Background: The filamentous fungus Trichoderma reesei is the main industrial cellulolytic enzyme producer. Several strains have been developed in the past using random mutagenesis, and despite impressive performance enhancements, the pressure for low-cost cellulases has stimulated continuous research in the field. In this context, comparative study of the lower and higher producer strains obtained through random mutagenesis using systems biology tools (genome and transcriptome sequencing) can shed light on the mechanisms of cellulase production and help identify genes linked to performance. Previously, our group published comparative genome sequencing of the lower and higher producer strains NG 14 and RUT C30. In this follow-up work, we examine how these mutations affect phenotype as regards the transcriptome and cultivation behaviour.

Results: We performed kinetic transcriptome analysis of the NG 14 and RUT C30 strains of early enzyme production induced by lactose using bioreactor cultivations close to an industrial cultivation regime. RUT C30 exhibited both earlier onset of protein production (3 h) and higher steady-state productivity. A rather small number of genes compared to previous studies were regulated (568), most of them being specific to the NG 14 strain (319). Clustering analysis highlighted similar behaviour for some functional categories and allowed us to distinguish between induction-related genes and productivity-related genes. Cross-comparison of our transcriptome data with previously identified mutations revealed that most genes from our dataset have not been mutated. Interestingly, the few mutated genes belong to the same clusters, suggesting that these clusters contain genes playing a role in strain performance.

Conclusions: This is the first kinetic analysis of a transcriptomic study carried out under conditions approaching industrial ones with two related strains of T. reesei showing distinctive cultivation behaviour. Our study sheds some light on some of the events occurring in these strains following induction by lactose. The fact that few regulated genes have been affected by mutagenesis suggests that the induction mechanism is essentially intact compared to that for the wild-type isolate QM6a and might be engineered for further improvement of T. reesei. Genes from two specific clusters might be potential targets for such genetic engineering.

No MeSH data available.


Related in: MedlinePlus